A mecA-containing Staphylococcus aureus strain was grown in the presence of high concentrations of D-serine, D-threonine, and D-phenylalanine. These growth conditions resulted in the replacement of the carboxyl-terminal (fifth) D-alanine residue of peptidoglycan stem peptides with the D-amino acid present in the growth medium and a reduced ability to grow in the presence of methicillin. The most dramatic effect was seen with D-serine. With 32 mM D-serine, strains that had been able to grow in the presence of 800 micro g of methicillin per ml were only able to grow in the presence of less than 50 micro g/ml. The results also suggest that in S. aureus vancomycin resistance mediated through the incorporation of precursors not terminating in D-alanyl-D-alanine would be mutually exclusive with expression of mecA-mediated methicillin resistance.